Impairment of eNOS contributes to the pathological alterations in vascular reactivity and structure that are observed in atherosclerosis. The premise of the current proposal is that impairment of the NOS pathway also plays a major role in the development of TA. We propose that in transplant recipients, the endothelial NOS pathway is impaired by two major insults: 1) increases in vascular superoxide anion (O2-), and 2) increases in ADMA. These abnormalities increase NO degradation and reduce NO synthesis. We further hypothesize that CMV infection augments these abnormalities via cytokine induced alterations in oxidative stress and ADMA accumulation. Accordingly, we propose to characterize the derangement of the NOS pathway in TA. We will assess the activity of the NOS pathway in transplant recipients using coronary vascular reactivity studies, coronary arteriovenous gradients and tissue biopsy studies using markers, which reflect NOS activity and oxidative stress. Genomic DNA obtained from the biopsy material will be used to characterize polymorphisms of the NOS pathway that may be associated with specific phenotypes. In addition, biochemical and physiological measurements of traditional and non-traditional risk factors, viral load and strain identification of CMV will be performed. These studies should provide insights into the mechanisms by which the NOS pathway is disturbed, and the role of CMV in these perturbations. However, cardiac transplant recipients have multiple metabolic disturbances that may impair the NOS pathway. Furthermore, CMV infection may have indirect effects on endothelial function by activation of the immune response. Therefore, to determine if there are direct effects of CMV infection on endothelial NOS, and to delineate mechanisms, additional studies of the CMV/NOS interaction will be conducted in endothelial cell cultures to determine the role of ADMA and oxidative stress in CMV-induced alterations in endothelial NOS and determinants of endothelial-mononuclear cell interaction.